A fluorescence quenching study of a sole tryptophan residue of a bifunctional alginate lyase from Pseudoalteromonas sp. strain No. 272 was done in the presence and absence of substrates, oligomeric guluronic and its C5 isomer mannuronic acid, by a Stern-Volmer plot with a quencher, acrylamide. N-Acetyltryptophanamide and reduced and carboxymethylated alginate lyase showed large quenching constants, on the other hand, the native enzyme had small constants regardless of the presence or absence of the substrates. The result suggests that the tryptophan residue is located in a buried region of the enzyme molecule, but is barely accessible to acrylamide, and that the residue is not masked by the substrates with various degrees of polymerization.